The interaction of the HDL protein apoA-l with the cell membrane transporter ABCA1 removes excess cellular cholesterol and protects against atherogenesis. This process is mediated by lipid-poor apoA- generated by either de novo synthesis or dissociation from HDL particles. Thus, factors that impair the availability or cholesterol efflux activity of apoA-l could have profound atherogenic effects. Oxidative damage is implicated in the pathogenesis of atherosclerosis, a chronic inflammatory disease. Oxidatively modified apoA-l have been detected in atherosclerotic lesions, and most of the apoA-l in HDL isolated from lesions has been structurally modified. We found that HOCI and acrolein, two common reactants generated by oxidation reactions, severely impair the ability of apoA-l to remove cellular cholesterol by the ABCA1 pathway and structurally modify the protein so as to generate large non-covalent complexes and amyloid-like fibrils. These results are consistent with the possibility that oxidation of apoA-l in vivo is atherogenic. We propose to test the hypothesis that HOCI and acrolein modify apoA-l by specific reactions so as to cause selective conformational switches that impair apolipoprotein function, and that these modifications contribute to the increased atherogenesis associated with inflammatory disorders. We will investigate the impact of HOCI and acrolein on the structure and function of apoA-l and small apolipoprotein-mimetic peptides, engineer apoA-l and mimetic peptides that are resistant to functional damage by HOCI and acrolein, and determine if oxidation-resistant apoA-l and mimetic peptides are atheroprotective in mouse models. This project will use mass spectrometric and physiochemical analyses to characterize structural changes in modified apoA-l and model peptides and in apoA-l isolated from atherosclerotic lesions, cell biology procedures to determine the effects of apoA-l modification on lipid transport activity and interactions with ABCA1, and mouse models to test for the atheroprotective effects of apoA-l and mimetic peptides engineered to be oxidation resistant. The proposed studies will provide insights into oxidation reactions that damage apoA-l in the artery wall and impair its atheroprotective function and help design oxidation-resistant peptides that can be used as therapeutic agents for treating cardiovascular disease.